Ovariectomy Group Research Articles

Inhibition of osteogenic and adipogenic potential in bone marrow-derived mesenchymal stem cells under osteoporosis

Osteoporosis, a prevalent systemic bone disease, has emerged as one of the most complicated health issues due to the risk of increased susceptibility to fractures. Bone-marrow mesenchymal stem cell (BMSC) has great potential of differentiating into several distinct cell types, including osteoblasts, adipocytes and chondrocytes. The present study analyzed the biological function changes of BMSCs under osteoporotic micro-environment and aimed to find a specific mechanism associated with this condition. Female rats were assigned to two groups: sham operation (SHAM) group and ovariectomy (OVX) group. BMSCs were harvested and cultured in vitro after 3 months post-ovariectomy. Alamar-Blue test suggested a higher proliferation ability in SHAM group. The differentiation potential of BMSCs was verified through various assays in vitro. RT-PCR and western blot analysis further confirmed the lower osteogenic and adipogenic differentiation capacity in OVX group. Moreover, through the microarray analysis, we were stunned to find that Integrin Alpha-7 (ITGA7) may improve osteogenesis through phosphatidylinositol 3-kinase/Akt (PI3K/Akt) signaling pathway. Overall, our study showed that osteoporosis inhibited the proliferation and differentiation of BMSCs, especially the osteogenesis and adipogenesis. Meanwhile, modulation of ITGA7 expression through PI3K/Akt signaling pathway might provide a new therapeutic target for osteoporosis.

Co-treatment of vitamin D supplementation and aerobic training improves memory deficit in ovariectomized rat

Objective: Metabolic syndrome (MetS) and insufficient vitamin D levels are globally increasing phenomena which are correlated with cognitive impairment. This study investigated the interactive effect of aerobic training with vitamin D supplementation on memory deficit in rats with metabolic syndrome induced by ovariectomy.Methods: A total of forty Wistar rats weighing 240–255 gr were randomly matched on their body weight and divided into ovariectomy (OVX, n = 32) and sham-operated (SHAM; n = 8) groups. OV group was then divided into vitamin D supplementation (OVX + Vit D; 10000 IU/kg/week, for 8 weeks, n = 8), aerobic training (OVX + AT; n = 8), aerobic training and vitamin D supplementation (OVX + AT + Vit D; 10000 IU/kg/week, for 8 weeks, n = 8), and vehicle control group receiving sesame oil (OVX + Ses Oil; n = 8). After the end of intervention, passive avoidance learning and memory were assessed in step through passive avoidance paradigm. Obtained data were analyzed by ANOVA and post hoc Tukey test.Results: After 8 weeks of aerobic training and vitamin D supplementation, step through dark compartment latency (STL) was significantly higher and total time spent in that compartment (TSD) was lower in OVX + AT + Vit D compared to the other counterpart groups.Conclusion: Vitamin D supplementation combined with 8-week aerobic training alleviates cognitive impairment metabolic syndrome induced by ovariectomy.

Genistein improves the vaginal epithelium thickness in a rat model of vaginal atrophy through modulation of hormone and heat shock protein 70 levels

Objective: To investigate whether administration of genistein can improve hormonal changes (estradiol and follicle stimulating hormone (FSH)] , heat shock protein 70 (Hsp70) expression, and thickness of vaginal epithelial cells in a rat model of vaginal atrophy. Methods: Twenty-five multiparous female rats were divided into five groups, namely the control group (without ovariectomy), the ovariectomy only group, and the ovariectomy groups 1, 2 and 3 receiving genistein at the doses of 0.045, 0.090 and 0.180 mg/kg body weight/day, respectively. Estradiol, FSH, and Hsp70 expression were analyzed by using the immunoassay technique. Analysis of the thickness of the vaginal epithelium was performed by histology. Results: Ovariectomy significantly decreased estradiol levels compared to the control group (P 0.05). Ovariectomy significantly increased parabasal cell Hsp70 expression compared to the control group (P 0.05). Conclusions: Genistein is able to increase the thickness of the vaginal epithelium through hormone modulation and cellular stress suppression. Genistein is beneficial in the form of a herbal or alternative food for improvement of vaginal atrophy due to menopause.

Subchondral bone dysplasia mediates susceptibility to osteoarthritis in female adult offspring rats induced by prenatal caffeine exposure

Our previous studies confirmed that prenatal caffeine exposure (PCE) could induce susceptibility to osteoarthritis in adult offspring rats due to poor chondrocyte differentiation, but its mechanism remains to be further investigated. This study aimed to explore whether subchondral bone dysplasia mediates susceptibility to osteoarthritis in adult offspring rats induced by PCE. Pregnant Wistar rats were treated with caffeine (120 mg/kg.d) or saline from gestational day (GD) 9 to 20. The female offspring were euthanized to collect femurs at GD20, postnatal week (PW) 6, and PW28 (non-ovariectomy and ovariectomy groups) to detect osteoarthritis-like phenotype, subchondral bone mass, ossification center development, and other evidence. The results showed that PCE increased the Mankin score of pathological articular cartilage, but decreased articular cartilage thickness and subchondral bone mass, which were more obvious after ovariectomy. Meanwhile, the correlation analysis results demonstrated that the Mankin score of articular cartilage was significantly negatively correlated with subchondral bone mass, and the thickness of articular cartilage was significantly positively correlated with subchondral bone mass. Further, the length and area of the primary and secondary ossification centers, the number of osteoblasts, and the related genes' expression of osteogenic differentiation (e.g., Runx2, BSP, ALP, and OCN) were all significantly decreased in the PCE group before and after birth. Taken together, PCE induced susceptibility to osteoarthritis in adult female offspring, which was likely related to the subchondral bone dysplasia and reduction of subchondral bone mass production due to developmental disorder of primary and secondary ossification centers caused by osteoblast differentiation disability before and after birth.

THE EFFECTIVENESS OF ZINC AND RISEDRONATE ON BONE TURNOVER IN RAT MODEL OF OSTEOPOROSIS ASSESSED WITH THE EXPRESSION OF β-CROSSLAPS

Background: Osteoporosis prevalence in Indonesia is high. Risedronate commonly used for treating the osteoporosis. Unfortunately, long term therapy can cause hazardous adverse effect to the patient. Previous research in rat model osteoporosis, Zinc have a potential effect to prevent and rebuild bone structure with osteoblast stimulation in bone formation and osteoclast activity inhibition in bone resorption by increasing Osteoprotegerin production, that inhibit bond between RANKL and RANK.Purpose: To determine the effectiveness of Zink and Risedronate on bone turnover in osteoporotic model rats assessed the β-CrossLaps expression.Methods: This research is conducted as experimental comparative method used 25 female Galur Wistar rat that divided into Non-ovariectomy and Ovariectomy group to induce osteoporosis. The treatment is divided into Zinc-Risedronate, and Zinc, or Risedronate only. One Way Anova and Independent T-test was used to analyzed the significant difference between groups.Result: One Way Anova statistic test forβ-CrossLaps expression value in the 3rd Taken blood sample showed that p is less than 0.05 (p<0.05), which indicated that are significant difference in the mean between variable at 3rd Taken blood sample in all group. Meanwhile, Independent T-test was used to compare β-CrossLaps expression in 3rd taken blood sample between Ovari and Ovari+Zinc, Ovari and Ovari+Zinc+Rise showed p is less than 0.05 (p<0.05), which indicated that there are significant difference in the mean between variable 2 groups.Conclusion: Zinc-Risedronate administration will decrease bone turnover and β-CrossLaps expression more than Zinc or Risedronate only in rat model osteoporosis.

Biochemical and immunohistochemical investigations on bone formation and remodelling in ovariectomised rats with tamoxifen citrate administration.

Osteoporosis results with the imbalance between osteoblastic formation and osteoclastic resorption, resulting in susceptibility to bone fractures. Ovariectomy leads to osteoporosis by triggering alterations in bone formation and structure. Tamoxifen as an anti-oestrogen is used for adjuvant therapy especially in metastatic diseases and known to have a bone mass protective effect after ovariectomy. An animal model of ovariectomy induced osteoporosis after tamoxifen citrate administration was studied via biochemical and immunohistochemical methods. Female Wistar albino rats (n = 45), selected according to their oestrous cycle, were divided into three groups; I - control, II - ovariectomy, III - ovariectomy + tamoxifen. Following ovariectomy, tamoxifen citrate (10 mg/kg) was given intraperitoneally daily for 8 weeks. At the end of the period, animals were sacrificed under anaesthesia, blood samples were taken to measure oestrogen, calcium, and alkaline phosphate. Tibia bone samples were fixed in formalin solution and decalcified with 5% ethylene-diamine tetra acetic acid. After the routine histological follow up, samples were embedded in paraffin and cut with a microtome for semi-thin sections. Primary antibodies osteonectin and osteopontin were applied to sections and examined under light microscope. As a consequence, when oestrogen and calcium data were compared there was a decrease in ovariectomy group with an increase in alkaline phosphatase. In ovariectomy + tamoxifen group, these values were close to the control group. Osteonectin was observed to promote bone formation by influencing collagen fibre formation, extracellular matrix development, osteoblast differentiation and the capacity to affect osteoclast activity. It has been suggested that osteopontin, the cytokine and cell binding protein, stimulates cellular signalling pathways, induces bone remodelling and acts in osteoporosis.

MiR-373 promotes the osteogenic differentiation of BMSCs from the estrogen deficiency induced osteoporosis.

This study aimed to investigate the expression of miR-373 in osteoporosis patients and rat models induced by estrogen deficiency and to detect whether miR-373 can regulate the ability of osteogenic differentiation of bone marrow mesenchymal stem cells in the osteoporosis microenvironment caused by estrogen deficiency. Bone tissues and blood samples were collected from 20 osteoporotic patients and 20 controls. PCR analysis was used to detect the expression of miR-373 in bone tissue and serum from postmenopausal osteoporotic patients and normal patients. 120 SD rats were purchased and randomly divided into sham operation group and OVX group. Rat models of sham-operated and bilateral oophorectomy mice models were constructed. The expression of miR-373 in bone tissue, cells, and serum of the mice was tested. Then, bone marrow mesenchymal stem cells from sham-operated rats and bilaterally ovariectomized rats were isolated and cultured. After 10 days of osteogenic induction, alkaline phosphatase staining and alizarin red staining were performed to test the osteogenic differentiation ability of bone marrow mesenchymal stem cells, and whether miR-373 affects this ability. PCR results showed that the expression of miR-373 in the bone tissue and the serum of patients with postmenopausal osteoporosis was significantly reduced. The expression of miR-373 was markedly decreased in the bone tissue, cells, and serum from the rats of bilateral ovariectomy group. Alkaline phosphatase staining and alizarin red staining showed that miR-373 could promote the differentiation of bone marrow mesenchymal stem cells into osteoblasts and reverse the decreased osteogenic differentiation of bone marrow mesenchymal stem cells caused by osteoporosis. The expression of miR-373 is decreased in osteoporotic patients and rat models caused by estrogen deficiency, and it can promote the differentiation of bone marrow mesenchymal stem cells into the osteogenic direction. This work provides a new direction and experimental basis for clinical diagnosis and treatment of osteoporosis.

Risedronate inhibits rat bone marrow adipogenesis and reduces RANKL expression in adipocytes

To investigate the effects of risedronate on bone marrow adipogenesis and the expression of the receptor activator of nuclear factor κB ligand (RANKL) in adipocytes in the bone marrow micro-environment. Primary cultured rat mesenchymal stem cells (BMSCs) with or without adipogenic induction for 14 days were treated with 1, 5, 10, and 25 μmol/L risedronate. The droplets of the differentiated adipocytes were analyzed, and Western blotting was performed to detect the expression level of RANKL. Female SD rats (24-week-old) were randomly divided into sham-operated group and ovariectomy (OVX) group, and 12 weeks after the operation, the OVX rats were further divided into control group and risedronate group (2.4 μg/kg, injected subcutaneously for 3 times a week). Eight weeks later, the bone mineral density (BMD) of the rats and bone marrow histopathology of the femurs was examined to evaluate the effect of risedronate on the fat fraction in the bone marrow. Risdronate significantly inhibited adipogenic differentiation of rat BMSCs and suppressed RANKL expression in the adipocytes derived from the BMSCs in a concentration-dependent manner. In OVX rats, risdronate treatment significantly increased the BMD and decreased the fat content in the bone marrow. Risdronate can effectively inhibit the adipogenic differentiation of rat BMSCs, decrease fat content in the bone marrow, and suppress the generation and function of osteoclasts by down-regulating the expression of RANKL, which can be an important mechanism underlying the therapeutic effect of risedronate against osteoporosis.

The Effects of Genistein on Renal Oxidative Stress and Inflammation of Ovariectomized Rats

Objectives: This study aimedatdetermining whetheran8-week genisteintreatment induced anti-inflammatoryand antioxidative stress effects in the uterine of Ovariectomized (OVX) Wistar rats. Methods: Three groups were formed including 10 female Wistar rats each: control (sham), ovariectomy group (OVX); OVX + genistein (1 mg/kg, daily SC for eight weeks). The genistein treatment was initiated 10 days after surgery and continued for two months. Renal malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase activity, tumor necrosis factor-α (TNF-α), and Interleukin-1β (IL-1β) were measured with Enzyme Linked Immunosorbent Assay (ELISA) and Western blot. Results: Higher amounts of MDA, TNF-α, and IL-1β were measured in the OVX group compared to the control group (P < 0.05), however, the amount of SOD, GPX, and catalase were significantly lower. The level of MDA, TNF-α, and IL-1β were significantly lower, and SOD, GPX, and catalase activity were on a higher level in OVX + genistein in comparison with the OVX group (P < 0.05). Also, histological observation showed some abnormality in kidney architecture, which was improved by genistein administration. Conclusions: Genistein is an appropriate treatment in attenuation of oxidative stress and inflammation in the kidneys of ovariec-tomized rats. Copyright © 2019, Jundishapur Journal of Natural Pharmaceutical Products.

Tissue Composition and Biomechanical Property Changes in the Vaginal Wall of Ovariectomized Young Rats.

Ideal animal models are needed to reflect the changes in the biochemical and biomechanical properties of the vagina that occur in pelvic organ prolapse (POP). In this study, we aimed to demonstrate the short and long-term effect of menopause on the biochemical and biomechanical properties of rat anterior vaginas. Here, Sprague-Dawley rats were bilaterally ovariectomized to induce menopause. Rats without ovariectomy served as the normal control group (n=12). The histology changes and the expression of collagen I, III, and a-SMA were assessed to indicate the biochemical changes in the vagina 2 weeks, 4 weeks, and 16 weeks after ovariectomy (n=6 for 2 and 4 weeks, n=12 for 16 weeks). Uniaxial biomechanical testing was conducted in the control group and ovariectomized rats 16 weeks after ovariectomy. Compared with the control group, the ovariectomy group showed a significant increase in the expression of collagen I 2 weeks after ovariectomy, while collagen III showed a declining trend. Two weeks after ovariectomy, the smooth muscle bundles began to become disorganized, and the fraction of smooth muscle in the nonvascular muscularis of the proximal vagina was significantly decreased (P<0.001). However, there was no difference in the expression of a-SMA in the distal vagina. Compared with the control group, the ovariectomy group had stiffer vaginas with a declining trend in the ultimate load 16 weeks after ovariectomy. In conclusion, surgically induced menopause had a significant short- and long-term impact on tissue composition and biomechanical properties of the rat vagina, which may lead to increased susceptibility to POP development.

Angiogenesis Changes in Ovariectomized Rats with Osteoporosis Treated with Estrogen Replacement Therapy.

To investigate whether angiogenesis changes in early menopausal osteoporosis treated with estrogen replacement therapy, 120 rats were randomly divided into five groups: sham operation group (SHAM), ovariectomy group (OVX), and ovariectomy plus three different estrogen doses replacement therapy groups (OVX + E2). We detected the bone microarchitecture and measured the expression levels of estrogen receptor beta (ERβ), vascular endothelial growth factor (VEGF), osteoprotegerin (OPG), and receptor activator of NF-κB ligand (RANKL). CD31 immunofluorescence and silica gel perfusion imaging were used to analyze the vascular distribution. We confirmed that the femur BMD of ovariectomized rats was significantly lower than SHAM group and OVX+E2 groups. After estrogen therapy, the local microvascular formation increased after estrogen treatment in a dose dependent manner. ERβ was downregulated and VEGF was upregulated, positively correlated with estrogen dosage. We successfully constructed an osteoporosis model of ovariectomized rats with estrogen replacement therapy. We also found angiogenesis changed in early menopausal osteoporosis treated with estrogen replacement therapy. We indicated that estrogen replacement therapy increased angiogenesis through VEGF upregulation. However, we observed that, at the highest doses of estrogen studied, increased angiogenesis was associated with a decrease in BMD, the underlying mechanisms of which remain unclear.

The effect of risedronate on orthodontic tooth movement in ovariectomized rats

ObjectiveThis study aimed to investigate the effect of risedronate on orthodontic tooth movement in ovariectomized rats. MethodsForty-five female Sprague-Dawley rats were divided into three groups. Bilateral ovariectomies were performed on the rats in the ovariectomy and ovariectomy + risedronate groups. Two weeks after ovariectomy, risedronate was administered intraperitoneally every three days in the ovariectomy + risedronate group, while rats in the ovariectomy and sham groups were administered saline until they were sacrificed. One month after ovariectomy, nickel-titanium alloy closed-coil springs were placed between the maxillary left first molar and the maxillary left incisor. On days 3, 7 and 14, five rats from each group were sacrificed. The distance of orthodontic tooth movement was measured by a digital caliper, and slides were obtained for histological analysis. ResultsThe distance of orthodontic tooth movement and the number of tartar-resistant acid phosphatase (TRAP)-positive cells were the highest in the ovariectomy group, followed by those in the ovariectomy + risedronate group, on days 3, 7 and 14. The positive expression levels of receptor activator of nuclear factor-kappa β (RANK) ligand and cathepsin K were the strongest while the positive expression of osteoprotegerin in the ovariectomy group was the weakest, followed by the corresponding expression levels in the ovariectomy + risedronate group, on days 3, 7 and 14. ConclusionsRisedronate can inhibit orthodontic tooth movement in ovariectomized rats and may function by regulating the RANK/RANK ligand/osteoprotegerin pathway.

Comparing the effect of tempe flour and tofu flour consumption on estrogen serum in ovariectomized rats

Estrogen decreasing during menopause can create problems in the cardiovascular organs, brain, urogental tract, and bone. Estrogen replacement therapy (ERT) can be used to increase estrogen levels. However, side-effects including breast cancer may limit their use. Tempe and tofu are natural plat-based foods which contain phytoestrogens. The aim of this research was to compare estrogen serum levels in ovariectomized rats given tempe flour and tofu flour.This research was conducted on female rats, aged 12 months (n = 72 rats). Rats were grouped into 5 groups: tempe flour, tofu flour, estradiol, casein and non-ovariectomized. The intervention was carried out for two months with three observation points; i) in the second week, ii) fifth week and iii) eighth week. Estrogen serum analysis was done by ELISA (Estradiol EIA-2693). The mean and the differences between treatment groups were analysed using one way ANOVA with post hoc polynomial contrasts (LSD).The highest estrogen serum in the second week intervention was found in the estradiol group followed by the tempe flour, tofu flour, non-ovariectomized and casein groups, respectively. The tempe flour group had the highest mean serum estrogen levels in the fifth week, followed by the estradiol group, non-ovariectomized group, tofu group and casein group. At the eighth week of intervention, the highest estrogen serum level was found in the tempe flour group followed by the estradiol group, tofu flour group, non ovariectomy group and casein group. Interventions in the fifth and eighth weeks showed significant differences between groups (p < 0.05). It was concluded that tempe flour rendered twice the serum estrogen level compared to tofu flour. Further research is needed in the form of clinical trials to prove that effect in humans.

Downregulation of Prdx2 Protects Osteoporosis Rats by Regulating Receptor Activator of Nuclear Factor Kappa-B/Osteoprotegerin Pathway

Osteoporosis (OP) is a bone disease caused by various causes and can be found in various stages, such as juvenile, adult, menopausal and old age. OP has the systemic, degenerative, and metabolic characteristics, which leads to loss of bone mass, structural changes, and biomechanical properties degeneration. Prdx2 is a member of the peroxiredoxase family with antioxidant effects and involved in the regulation of bone and joint diseases. However, the role of Prdx2 in OP and related mechanisms has not been elucidated. SD rats were randomly divided into 2 groups, OP group in which OP rat model was prepared by ovariectomy and sham operation group. Prdx2 siRNA was transfected into OP rats followed by analysis of the expression of Prdx2, Opn, and Osterix by real-time PCR, bone density changes by dual energy line bone densitometer, formation of mineralized nodules by Alizarin red staining, Serum osteocalcin (OC) activity by ELISA as well as RANK and osteoprotegerin (OPG) expressions by real-time PCR and Western blot. Prdx2 expression was significantly increased, bone mineral density (BMD) was reduced, mineralized nodule formation was attenuated, Opn and Osterix levels were downregulated, together with decreased serum OC activity, increased RANK expression, and declined OPG expression apparently in OP rats compared with sham group (P &lt; 0.05). Prdx2 siRNA transfection downregulated Prdx2 and RANK levels, increased BMD, mineralized nodule formation, Opn, OPG, and Osterix levels, as well as serum OC activity in OP rats (P &lt; 0.05). Prdx2 expression is elevated in osteoporotic rats, which is associated with decreased osteogenic differentiation and BMD, leading to osteoporosis. Downregulation of Prdx2 expression can improve osteoporosis by regulating the RANK/OPG pathway.

Estrogen Deficiency Exacerbates Intervertebral Disc Degeneration Induced by Spinal Instability in Rats.

An experimental animal study of osteoporosis (OP) and intervertebral disc degeneration (IDD). The aim of this study was to clarify the effects of estrogen deficiency and supplement on cervical IDD induced by bilateral facetectomy in rats. The relationship between IDD and OP is still controversy with the wide prevalence in aged people. Seventy-two Sprague-Dawley female rats were randomly divided into ovariectomy (OVX) group, facet joints resection of C4-6 (FR), FR-OVX group, estrogen replacement therapy (ERT, based on the FR-OVX group) group, and sham group. Specimens of C4-6 segment were harvested at 12 and 24 weeks. The microstructures of C5 vertebrae, vertebral endplate lesions and calcification, and IDD of C5/6 disc were evaluated by micro-computed tomography (micro-CT) and histology. The protein and gene levels of aggrecan, Col2α1, matrix metalloprotease (MMP)-3, and MMP-13 in the C5/6 and C4/5 discs were measured. Microstructures of C5 vertebral body were weakened significantly after ovariectomy, while restored effectively with estradiol supplementation. The facetectomy led to significant IDD, and the IDD was aggravated when combined with OVX. The IDD of the ERT group was alleviated effectively and similar to that of the FR group in intervertebral disc height, vertebral endplate lesions and calcification, and disc degeneration scores. In addition, the estrogen supplement maintained the extracellular matrix by decreasing MMP-3 and MMP-13, and increasing aggrecan and Col2α1 expression. The present study demonstrated that estrogen deficiency exacerbated IDD induced by spinal instability, while estrogen supplementation alleviated the progression of disc degeneration related to osteoporosis. N/A.

Effect of moxibustion at "Guanyuan" (CV 4) and "Sanyinjiao" (SP 6) on bone morphology, metabolism and ERα of bone marrow mesenchymal stem cells in the ovariectomized rats

To observe the effects of moxibustion at "Guanyuan" (CV 4) and "Sanyinjiao" (SP 6) on bone morphology, metabolism and ERα of bone marrow mesenchymal stem cells (MSCs) in the ovariectomized rats and explore the underlying mechanism of moxibustion at Guanyuan (CV 4) and Sanyinjiao (SP 6) on the regulation of bone metabolism. A total of 60 SD rats were randomized into a normal group (20 rats) and an ovariectomy group (40 rats). In the normal group, no any treatment was provided. In the ovariectomy group, the classical ovariectomy was adopted to set up the osteoporosis model. In the 13th week after ovariectomy, 10 rats were collected from each of the normal group and the ovariectomy group for model identification. After model identification, the ovariectomy group was subdivided into a model group, an estradiol group and a moxibustion group, 10 rats in each one. In the normal group and the model group, the gavage was provided with 2 mL 0.9% sodium chloride solution, once a day. In the estradiol group, the gavage was provided with 17β-estradiol 100 μg/(kg•d), once a day. In the moxibustion group, moxibustion was applied at "Guanyuan" (CV 4) and "Sanyinjiao" (SP 6), 15 min at each acupoint, once a day. The 6 treatments were as one course, with 1 day of interval, 12 courses were required. After 12-week intervention, the dual-energy X-ray was adopted to determine the bone mineral density (BMD) and bone mineral content of the whole body in living condition. After sacrificed, HE staining was adopted to observe femur structure, the enzyme-linked immunosorbent assay was to determine the absorbance of estradiol (E2) and alkaline phosphatase (ALP) in serum and the real-time quantitative PCR method was to determine the mRNA expressions of ERα of MSCs in tibia and femur. Compared with the model group, BMD was increased obviously in the pelvis and spine in the moxibustion group and the estradiol group (P<0.05, P<0.01). Compared with the model group, the bone mineral content was higher in the rib, pelvis and spine in the moxibustion group and the estradiol group, there was no statistically significant difference (all P>0.05). Compared with the model group, the bone trabeculas were stronger and well connected in the moxibustion group, indicated by HE staining. Compared with the model group, the concentrations of E2 in serum were increased obviously in the estradiol group and the moxibusiton group (both P<0.01), and ALP concentrations reduced obviously in the estradiol group and the moxibustion group (both P<0.001), the mRNA expressions of ERα in MSCs increased in the estradiol group and the moxibustion group (both P<0.001). Moxibustion at "Guanyuan" (CV 4) and "Sanyinjiao" (SP 6) effectively increases BMD and bone strength in the ovariectomized rats and the mechanism may be related to the improvement of serum E2 concentration, the decrease of serum ALP concentration and the up-regulation of mRNA expression of ERα in MSCs.

Effect of Bushen Jiangu decoction on ovariectomyinduced osteoporosis in rats

Purpose: This study aimed to determine the therapeutic effect of Bushen Jiangu Decoction (BJD) on osteoporosis (OS) in rats.Methods: The animals were divided into the following groups: control group, rats received no treatment; ovariectomy (OVX) group, rats subjected to OVX surgery and were treated with normal saline; OVX + Fosamax group, rats that received OVX and were treated with 2 mg/kg/week of Fosamax; low-dose BJD (40 mg/kg/day), medium-dose BJD (80 mg/kg/day) and high-dose BJD (160 mg/kg/day) groups, where rats received OVX surgery with BJD doses of 40, 80 and 160 mg/kg/day, respectively. At four weeks after treatment with OVX, Fosamax or BJD was orally administered for four months. Bone mineral density (BMD) was assessed, while serum hormone, serum levels of follicle-stimulating hormone (FSH), alkaline phosphatase (ALP), estradiol (E2), luteinizing hormone (LH), osteocalcin (OC) and telopeptides of collagen type I (CTx) were measured by enzyme linked immunosorbent assay (ELISA).Results: The results demonstrated that the reduced BMDs of L4 femurs and vertebrae were inhibited by BJD. Furthermore, BJD significantly elevated the serum levels of FSH, E2 and LH in OS rats. Furthermore, the serum CTx, ALP and OC levels of these rats dramatically decreased, when compared to the OVX group.Conclusion: These findings suggest that BJD can improve OVX-induced OS.Keywords: Bushen Jiangu decoction, Osteoporosis, Bone mineral density, Serum biochemistry

SCD1 methylation in subcutaneous adipose tissue associated with menopausal age

Objective: This study was designed to investigate associations between menopausal age and the DNA methylation levels of stearoyl-coenzyme A desaturase 1 (SCD1, selected by the Illumina Human Methylation 450 K Bead Chip) and explore the changes in mRNA levels of SCD1 and DNA methyltransferases (DNMTs) in response to estrogen replacement therapy (ERT).Methods: In the human experiment, we performed subcutaneous adipose tissue DNA extraction on 85 menopausal women. Methylation of SCD1 was measured by MethyLight polymerase chain reaction. In the rat experiment, we established models of menopause (ovariectomy group) and ERT (ovariectomy + 17β-estradiol group). The mRNA levels of SCD1, DNMT1, DNMT3A, and DNMT3B were determined.Results: The results showed that DNA methylation of SCD1 was inversely correlated with menopausal age (r = 0.370, P < 0.001). In the rat study, the mRNA levels of SCD1 decreased (P < 0.001) and those of DNMT3A (P < 0.001) and DNMT3B increased (P < 0.001) after ERT.Conclusion: Methylation levels of SCD1 were significantly associated with menopausal age. DNMT3A and DNMT3B may be involved in the methylation of SCD1.

The effect of low-level estrogen in mandibular bone: An in vivo study

Low levels of estrogen can cause osteoporosis and usually occur during a woman's menopausal phase. Osteoporosis can lead to bone resorption, the absence of osseointegration, and implant failure. The aim of this study is to determine the expression of transforming growth factor-beta 1 (TGF-β1), runt-related transcription factor (RUNX2), and osteoblasts in mandibular rats with low levels of estrogen. This study is an in vivo experimental research. Female Wistar rats (n = 18) were divided into two groups: (1) Postsham surgery and (2) ovariectomy group. After 12 weeks, the rats were sacrificed to identify the level of estrogen, while histological analysis was conducted to determine the level of osteoblast and the expression of TGF-β1 and RUNX2. The data were analyzed using t-test (P < 0.05). There were significant lower levels of estrogen and osteoblast among the ovariectomy group compared to the postsham group (P < 0.05). RUNX2 levels were found to be significantly higher in the ovariectomy group than that in the postsham group (P < 0.05). However, there were no significant differences between TGF-β1 levels within the ovariectomy and postsham groups (P > 0.05). Ovariectomy can lead to decreased osteoblastogenesis in mandibular bone by the reduced level of osteoblast and the increased expression of TGF-β1 and RUNX2.

Long-Term Nikotine Administration Impairs Blood Parameters and Femoral Bone Tissue Structure in an Osteoporosis Model of Rats

Osteoporosis is a systemic bone disease that results in the loss of bone mass and impared bone structure. Animal models for osteoporosis are generated by ovariectomy. Adult female Sprague-Dawley rats were divided into control, bilateral ovariectomy and bilateral ovariectomy and subcutaneous nicotine administered (received nicotine sulphate, 2 mg/kg) groups, daily for 28 days. At the end of the period, rats were sacrified under anesthesia blood samples were taken and femoral tissues were dissected. Estrogen, calcium and alkaline phosphatase levels were measured. Tissue samples were prepared for histopathogical examination. Sections were stained with Hematoxylin and Eosin and examined under light microcope. Biochemical parameters were decreased depending on the overiectomy, also decrement was notable with nicotine intake. In the ovariectomy group; increased inflammatory cells with degenerative changes around the femoral compact bone and dilatation of osteon structures in bone trabeculae and apoptotic changes in osteocyte cells in bone lacuna were apparent. In the ovariectomy with nicotine administration group, excessive dilatation of the havers lamellae in the compact bone region, increased osteoclastic activity, picnosis and apoptotic nucleus of the osteoclastic cells located in the lacuna, and increased collagen fibers in the matrix were observed. We suggest that ovariectomy and nicotine administration together effect estrogen and calcium metabolism negatively, stimulate alterations in the structural properties of bone matrix, also affect osteocyte development and bone lamellar structure that may accelerate osteoporosis development.